An In Vitro Method to Generate Astrocyte Scaffolds within Hydrogel Micro-columns

Take a culture dish containing a micro-column boat comprising hollow tubes of defined diameter.      

The micro-columns, made of a hydrogel, are filled with a buffer to prevent drying.

Under a stereomicroscope, remove the buffer and add an ice-cold collagen solution, a component of the extracellular matrix.   

Introduce a media along the dish's inner edge to maintain column hydration. Incubate under physiological conditions for the collagen to polymerize, forming a coating inside the micro-columns.         

Add astrocytes inside the micro-columns and incubate to promote astrocyte adhesion to collagen.

Next, fill the dish with the medium and maintain it under physiological conditions.       

Due to the micro-columns' narrow diameter, the astrocytes acquire a bipolar morphology with processes extending along the length.

Over time, the astrocytes self-assemble to form a three-dimensional scaffold containing dense bundles of astrocyte processes.

The hydrogel-encased astrocyte scaffold is ready for downstream assays.       

Inside a biosafety cabinet, prepare a 1 milligram per milliliter solution of rat tail type I collagen in co-culture medium and then place it on ice. Adjust the pH of